Unfolding and inactivation of abalone (Haliotis diversicolor) alkaline phosphatase during denaturation by guanidine hydrochloride.
Abalone, a kind of low poikilothermic invertebrate, is easily exposed to ocean environment stress. Since it is one of the important mariculture animals, the attention paid to the abalone study becomes increasing. Alkaline phosphatase (ALPase, EC 3.1.3.1) is a kind of zinc-contained metalloenzyme, which catalyzes the nonspecific hydrolysis of phosphate monoesters. Unfolding and inactivation of ALPase from abalone (Haliotis diversicolor) during denaturation by guanidine hydrochloride (GuHCl) of different concentrations has first been studied. The kinetic theory of the substrate reaction by enzyme was described by Tsou, which was applied to the study on ALPase's kinetic course of inactivation by GuHCl. The result showed that the inactivation of the enzyme by GuHCl was a slow, reversible reaction with fractional remaining activity. The microscopic rate constants were determined. The result, k(+0) > k'(+0), showed that the enzyme was protected by the substrate to a certain extent during guanidine denaturation. The changes of conformation of the enzyme in different concentrations of GuHCl have been studied by means of measuring the fluorescence spectra. The results showed that the inactivation occurred before the noticeable conformational changes of the enzyme molecule as a whole can be detected, which suggests that the active site of the enzyme has more flexibility than the whole enzyme molecule. These studies will facilitate the understanding of physiological and biochemical features of the H. diversicolor and will also help in the understanding of the abalone immune system.
Liao
Jin Hua
JH
Key Laboratory of Ministry of Education for Cell Biology and Tumor Cell Engineering, Xiamen University, Xiamen 361005, China.
Chen
Qing Xi
QX
Zhang
Qian
Q
Yang
Yi
Y
Shi
Yan
Y
D000469:Alkaline Phosphatase / Q000737:chemistry* / Q000378:metabolism*; D000818:Animals; D049851:Gastropoda / Q000201:enzymology*; D019791:Guanidine / Q000737:chemistry*; D011489:Protein Denaturation; D017510:Protein Folding
D000469:Alkaline Phosphatase; D019791:Guanidine